Capsular Exopolysaccharide Research Articles

Anti-quorum sensing activity of essential oils against multidrug-resistant Klebsiella pneumoniae: a novel approach to control bacterial virulence.

The increasing resistance of microbes to conventional drugs is a serious problem worldwide that has increased the need for alternative antimicrobial compounds. Naturally occurring essential oils (EOs) are considered an important component of traditional pharmacopeia because of their antimicrobial and antioxidant properties. This has attracted researchers to identify novel therapeutic anti-pathogenic agents that could act as non-toxic quorum sensing inhibitors, thus controlling infections without encouraging the development of bacterial resistance. This prompted to undertake the current investigation to unravel the efficacy of EOs as QS modulators in reducing the virulence of multidrug resistant (MDR) strains ofKlebsiella pneumoniae. The study highlighted the anti-QS activity of fifteen EOs in modulating the QS-related traits by a reduction in capsular polysaccharide, exopolysaccharide and siderophore production in addition to inhibition of biofilm formation. The overall results suggest using EOs to develop alternate intervention strategies to mitigate infections caused by MDR strains ofK. pneumoniae.

Investigating the impact of exopolysaccharides on yogurt network mechanics and syneresis through quantitative microstructural analysis

Exopolysaccharides produced by lactic acid bacteria are widely used to improve the sensory properties of yogurt. The relation between the physical properties of the microbial exopolysaccharides and the structural and rheological properties of the yogurt are incompletely understood to date. To address this knowledge gap, we studied how two distinct exopolysaccharides influence the microstructure, rheological properties, and syneresis of yogurt. The effect of a negatively charged, capsular exopolysaccharide produced by Streptococcus thermophilus and a neutral, non-capsular exopolysaccharide produced by Lactococcus lactis were investigated. Using quantitative microstructural analysis, we examined yogurt samples prepared with either the capsular or the non-capsular exopolysaccharide, and with mixtures of the two. Confocal laser scanning microscopy and stimulated emission depletion microscopy were employed to visualize the microstructures, revealing differences in pore size distribution, protein domain size, and casein interconnectivity that were not apparent through visual inspection alone. Additionally, variations in rheological properties were observed among the different yogurt types. In the yogurt fermented with both bacterial strains, we observed a combined impact of the two exopolysaccharide types on relevant microstructural and rheological properties. The negatively charged capsular exopolysaccharide enhanced casein interconnectivity and gel stiffness, while the neutral non-capsular exopolysaccharide led to thicker protein domains, an abundance of small pores, and a lower loss tangent. These factors collectively hindered syneresis, resulting in improved structural integrity. Our study not only provides valuable insights into the influence of different exopolysaccharides on yogurt properties, but also presents the first demonstration and quantification of the effect of multiple types of exopolysaccharides on casein interconnectivity. These findings offer guidance for the production of yogurts with customized microstructure, rheological properties, and resistance to syneresis.

Structural characteristics and functional properties of a fucose containing prebiotic exopolysaccharide from Bifidobacterium breve NCIM 5671.

To evaluate the structure and functions of capsular exopolysaccharide (CPS) from Bifidobacterium breve NCIM 5671. A CPS produced by the probiotic bacteria B. breve NCIM 5671 was isolated and subjected to characterization through GC analysis, which indicated the presence of rhamnose, fucose, galactose, and glucose in a molar ratio of 3:1:5:3. The average molecular weight of the CPS was determined to be ∼8.5×105 Da. Further, NMR analysis revealed the probable CPS structure to be composed of major branched tetra- and penta-saccharide units alternately repeating and having both α- and β-configuration sugar residues. CPS displayed an encouraging prebiotic score for some of the studied probiotic bacteria. Compared to standard inulin, CPS showed better resistance to digestibility against human GI tract in vitro. DPPH, total antioxidant, and ferric reducing assays carried out for CPS displayed decent antioxidant activity too. This study indicates that the CPS from B. breve NCIM 5671 has the potential to be utilized as a prebiotic food supplement. It is a high-molecular-weight (∼8.5×105 Da) capsular heteropolysaccharide containing rhamnose, fucose, galactose, and glucose.

Prospects of a hot spring-originated novel cyanobacterium, Scytonema ambikapurensis, for wastewater treatment and exopolysaccharide-enriched biomass production.

The present work performs the polyphasic characterization of a novel cyanobacterial species Scytonema ambikapurensis isolated from an Indian hot spring and evaluates its wastewater bioremediation potential. While the physicochemical analyses of the wastewater indicated high load of nutrients and metals, the wastewater bioremediation experiment performed using the test cyanobacterium denoted the removal of 70 and 86% phosphate, 49 and 66% sulfate, 96 and 98% nitrate, 91 and 92% nitrite, 95 and 96% ammonia, 66 and 72% chloride, 79 and 81% zinc, 68 and 80% nickel, 81 and 90% calcium, and 80 and 90% potassium from the autoclaved and un-autoclaved wastewater, respectively, after 20days of culturing. The kinetics study of zinc and nickel removal from wastewater revealed that the cyanobacterium employed sequential biosorption (by following pseudo-second-order kinetics model) and bioaccumulation methods to remove these two metals. The quality of the autoclaved and un-autoclaved wastewater was further improved by the cyanobacterium through reduction of hardness by 74 and 81%, respectively. In wastewater, the cyanobacterium not only enhanced its biomass, chlorophyll and carbohydrate contents, but also produced small amount of released and high capsular exopolysaccharide (EPS). The FTIR and TGA analyses of capsular EPS unraveled that it was a negatively charged sulfated biomolecule having thermostability up to 240°C, which suggested its possible use as excellent emulsifying, viscosifying, and biosorption agent. The credibility of this EPS as biosorption agent was ascertained by evaluating its metal chelating ability. Finally, the experimental data denoting the ability of S. ambikapurensis to bioremediate wastewater and simultaneously produce EPS was statistically validated by PCA1-pollutant removal model and the PCA2-cellular constituent model, respectively. Briefly, the study discloses that the cyanobacterium has huge biotechnological and industrial importance as it bioremediates wastewater and simultaneously produces thermostable exopolysaccharide.

Interactions between host epithelial cells and Acinetobacter baumannii promote the emergence of highly antibiotic resistant and highly mucoid strains

ABSTRACT Acinetobacter baumannii is an important nosocomial pathogen. Upon colonizing a host, A. baumannii are subjected to selective pressure by immune defenses as they adapt to the host environment. However, the mechanism of this pathoadaptation is unknown. Here, we established an in vitro system to evolve A. baumannii driven by the continuous selective pressure exerted by epithelial cells, and we used a combination of experimental evolution, phenotypic characterization and multi-omics analysis to address the underlying mechanism. When continuously exposed to selective pressure by pulmonary epithelial cells, A. baumannii showed ptk mutation-mediated mucoid conversion (reduced adhesion and increased anti-phagocytic ability) by enhancement of capsular exopolysaccharide chain length; rsmG mutation-mediated deficiency of 7-methylguanosine modification in the 524th nucleotide of 16S rRNA, which increased ribosome translation efficiency; and rnaseI mutation-mediated changes in outer membrane permeability and efflux pump expression. Together, these mutations altered susceptibility to a variety of antimicrobial agents, including the novel antibiotic cefiderocol, by regulating siderophore and siderophore-receptor biosynthesis. In conclusion, pulmonary epithelial cells modulate A. baumannii pathoadaptation, implicating the host–microbe interaction in the survival and persistence of A. baumannii.

Production of colanic acid hydrolysate and its use in the production of fucosylated oligosaccharides by engineered Saccharomyces cerevisiae

Fucose is a rare sugar obtained from fucoidan in brown algae. However, fucoidan contains low amounts of fucose, making the process of obtaining fucose inefficient and expensive. Colanic acid (CA) is a capsular exopolysaccharide produced by Escherichia coli, which has great potential as a source of fucose owing to its high proportion of fucose. In this study, we aimed to efficiently obtain fucose from the hydrolysis of CA and then produce 2′-fucosyllactose (2′-FL) and 2′-fucosylgalactose (2′-FG) using fucose in the CA hydrolysate, using engineered yeast. For the acid hydrolysis of CA, optimal conditions were 110 °C, 4% (w/v) H3PO4, and 60 min. Using fucose from CA hydrolysate as a substrate, 2′-FL and 2′-FG were produced from engineered Saccharomyces cerevisiae. This is the first report of producing fucosylated oligosaccharides from CA. Considering the benefits of CA in obtaining fucose, these results can lead to further applications of CA as a fucose supplier.

Characterisation of recombinant GH 3 β-glucosidase from β-glucan producing Levilactobacillus brevis TMW 1.2112

Levilactobacillus (L.) brevis TMW 1.2112 is an isolate from wheat beer that produces O2-substituted (1,3)-β-D-glucan, a capsular exopolysaccharide (EPS) from activated sugar nucleotide precursors by use of a glycosyltransferase. Within the genome sequence of L. brevis TMW 1.2112 enzymes of the glycoside hydrolases families were identified. Glycoside hydrolases (GH) are carbohydrate-active enzymes, able to hydrolyse glycosidic bonds. The enzyme β-glucosidase BglB (AZI09_02170) was heterologous expressed in Escherichia coli BL21. BglB has a monomeric structure of 83.5 kDa and is a member of the glycoside hydrolase family 3 (GH 3) which strongly favoured substrates with β-glycosidic bonds. Km was 0.22 mM for pNP β-D-glucopyranoside demonstrating a high affinity of the recombinant enzyme for the substrate. Enzymes able to degrade the (1,3)-β-D-glucan of L. brevis TMW 1.2112 have not yet been described. However, BglB showed only a low hydrolytic activity towards the EPS, which was measured by means of the D-glucose releases. Besides, characterised GH 3 β-glucosidases from various lactic acid bacteria (LAB) were phylogenetically analysed to identify connections in terms of enzymatic activity and β-glucan formation. This revealed that the family of GH 3 β-glucosidases of LABs comprises most likely exo-active enzymes which are not directly associated with the ability of these LAB to produce EPS.

Nanobiotechnological prospects of probiotic microflora: Synthesis, mechanism, and applications

Nanotechnology-driven solutions have almost touched every aspect of life, such as therapeutics, cosmetics, agriculture, and the environment. Physical and chemical methods for the synthesis of nanoparticles involve hazardous reaction conditions and toxic reducing as well as stabilizing agents. Hence, environmentally benign green routes are preferred to synthesize nanoparticles with tunable size and shape. Bacteria, fungi, algae, and medicinal plants are employed to synthesize gold, silver, copper, zinc, and other nanoparticles. However, very little literature is available on exploring probiotic bacteria for the synthesis of nanoparticles. In view of the background, this review gives the most comprehensive report on the nanobiotechnological potential of probiotic bacteria like Bacillus licheniformis, Bifidobacterium animalis, Brevibacterium linens, Lactobacillus acidophilus, Lactobacillus casei, and others for the synthesis of gold (AuNPs), selenium (SeNPs), silver (AgNPs), platinum (PtNPs), tellurium nanoparticles (TeNPs), zinc oxide (ZnONPs), copper oxide (CuONPs), iron oxide (Fe3O4NPs), and titanium oxide nanoparticles (TiO2NPs). Both intracellular and extracellular synthesis are involved as potential routes for biofabrication of polydispersed nanoparticles that are spherical, rod, or hexagonal in shape. Capsular exopolysaccharide associated carbohydrates such as galactose, glucose, mannose, and rhamnose, cell membrane-associated diglycosyldiacylglycerol (DGDG), 1,2-di-O-acyl-3-O-[O-α-D-galactopyranosyl-(1 → 2)-α-d-glucopyranosyl]glycerol, triglycosyl diacylglycerol (TGDG), NADH-dependent enzymes, amino acids such as cysteine, tyrosine, and tryptophan, S-layer proteins (SLP), lacto-N-triose, and lactic acid play a significant role in synthesis and stabilization of the nanoparticles. The biogenic nanoparticles can be recovered by rational treatment with sodium dodecyl sulfate (SDS) and/or sodium hydroxide (NaOH). Eventually, diverse applications like antibacterial, antifungal, anticancer, antioxidant, and other associated activities of the bacteriogenic nanoparticles are also elaborated. Being more biocompatible and effective, probiotic-generated nanoparticles can be explored as novel nutraceuticals for their ability to ensure sustained release and bioavailability of the loaded bioactive ingredients for diagnosis, targeted drug delivery, and therapy.

The Specific Capsule Depolymerase of Phage PMK34 Sensitizes Acinetobacter baumannii to Serum Killing.

The rising antimicrobial resistance is particularly alarming for Acinetobacter baumannii, calling for the discovery and evaluation of alternatives to treat A. baumannii infections. Some bacteriophages produce a structural protein that depolymerizes capsular exopolysaccharide. Such purified depolymerases are considered as novel antivirulence compounds. We identified and characterized a depolymerase (DpoMK34) from Acinetobacter phage vB_AbaP_PMK34 active against the clinical isolate A. baumannii MK34. In silico analysis reveals a modular protein displaying a conserved N-terminal domain for anchoring to the phage tail, and variable central and C-terminal domains for enzymatic activity and specificity. AlphaFold-Multimer predicts a trimeric protein adopting an elongated structure due to a long α-helix, an enzymatic β-helix domain and a hypervariable 4 amino acid hotspot in the most ultimate loop of the C-terminal domain. In contrast to the tail fiber of phage T3, this hypervariable hotspot appears unrelated with the primary receptor. The functional characterization of DpoMK34 revealed a mesophilic enzyme active up to 50 °C across a wide pH range (4 to 11) and specific for the capsule of A. baumannii MK34. Enzymatic degradation of the A. baumannii MK34 capsule causes a significant drop in phage adsorption from 95% to 9% after 5 min. Although lacking intrinsic antibacterial activity, DpoMK34 renders A. baumannii MK34 fully susceptible to serum killing in a serum concentration dependent manner. Unlike phage PMK34, DpoMK34 does not easily select for resistant mutants either against PMK34 or itself. In sum, DpoMK34 is a potential antivirulence compound that can be included in a depolymerase cocktail to control difficult to treat A. baumannii infections.

How centrifugation influences the recovery and soy peptone incorporation in hyaluronic acid coils from fermentation

Hyaluronic acid from fermentation (bio-HA) is a high-added-value biopolymer with a wide range of applications in aesthetic and medical areas. Bio-HA is a capsular exopolysaccharide highly hydrophilic with a coiled structure and high negative charge density at physiological pH. Centrifugation, currently used to separate microbial cells after fermentation, should significantly affect the bio-HA interactions with the medium components. Although the literature shows various centrifugation conditions for bio-HA, there is no concern about its recovery and characterization. We examined how centrifugation influences its recovery and characterized interactions with soy peptone (SP), used as a nitrogen source. We collected experimental data according to a statistical central composite design (CCD) planning with replicates at the central point with time (8–22 min) and spin (582xg to 12,402xg) as independent variables. The obtained results show bio-HA recoveries ranged from 45 ± 0.5–100% as dosed in the supernatant. The incorporation of amino acids and peptides from SP increased HA coils' density, causing losses due to sedimentation at higher spins and times. Furthermore, SP incorporation also changed the coils' size regarding HA standards of molar mass 106,105 and 104 Da. We conclude centrifugation should significantly impact downstream processing and the total cost of bio-HA production.

Antibiotic Modulation of Capsular Exopolysaccharide in Pelagicoccus enzymogenes sp. nov. Isolated From Marine Sediment

Although Verrucomicrobia is widely distributed in the marine environment, their physiological or cellular properties are poorly characterized because of the lack of cultured representatives. Under the selective pressure of two antibiotics, ofloxacin and norfloxacin, a Gram-stain-negative, aerobic coccus with exopolysaccharide (EPS) production ability was isolated from the coastal sediment of Xiaoshi Island, Weihai, China. These antibiotics inhibited bacterial growth, giving rise to the relatively slow-growing Verrucomicrobia that formed colonies on the isolation plates. It may be an effective method for the isolation of Puniceicoccaceae. From the taxonomic data obtained in this study, the new marine isolate NFK12T (=KCTC 72940T = MCCC 1H00424T) is proposed to be placed into a novel species within the genus Pelagicoccus for which the name Pelagicoccus enzymogenes sp. nov. is proposed. The EPS production of the strain NFK12T and the related strains were investigated and the effect of EPS produced by the strain NFK12T on the growth of other strains was examined. Besides, the effect of EPS on tolerance to ofloxacin and norfloxacin of the strain NFK12T was studied by measuring the biomass of the strain NFK12T. It was deduced that those strains that produced EPS tentatively protected themselves against the inhibitory effects of ofloxacin and norfloxacin.

Assembly of Bacterial Capsular Polysaccharides and Exopolysaccharides.

Polysaccharides are dominant features of most bacterial surfaces and are displayed in different formats. Many bacteria produce abundant long-chain capsular polysaccharides, which can maintain a strong association and form a capsule structure enveloping the cell and/or take the form of exopolysaccharides that are mostly secreted into the immediate environment. These polymers afford the producing bacteria protection from a wide range of physical, chemical, and biological stresses, support biofilms, and play critical roles in interactions between bacteria and their immediate environments. Their biological and physical properties also drive a variety of industrial and biomedical applications. Despite the immense variation in capsular polysaccharide and exopolysaccharide structures, patterns are evident in strategies used for their assembly and export. This review describes recent advances in understanding those strategies, based on a wealth of biochemical investigations of select prototypes, supported by complementary insight from expanding structural biology initiatives. This provides a framework to identify and distinguish new systems emanating from genomic studies.

Test parameters and cell chain length of Streptococcus thermophilus affect the microbial adhesion to hydrocarbons assay: a methodical approach.

The microbial adhesion to hydrocarbons (MATH) test is one of the most common method to determine the hydrophobicity of cell surfaces. Despite its prevalence, no standard test parameters are used in literature, making a direct comparison of data almost impossible. Criticism also focuses on test parameters that may mask hydrophobic interactions and hence lead to erroneous test results. We methodically investigated the impact of different MATH test parameters on the calculation of the cell surface hydrophobicity of Streptococcus thermophilus, a widespread exopolysaccharide-producing lactic acid bacterium used in the production of fermented milk products. Besides composition and ionic strength of the buffer used for cell re-suspension, we observed a pronounced time dependency of the turbidity of the cell suspension during phase separation due to sedimentation and/or cell lysis. A new modification of the MATH assay was applied to enable the determination of cell surface hydrophobicity of long chain-forming bacteria. As the cell surface hydrophobicity was not altered during exponential growth phase, we assume that the cell surface and its capsular exopolysaccharide layer are not changed during cultivation.

Genomic traits of Klebsiella oxytoca DSM 29614, an uncommon metal-nanoparticle producer strain isolated from acid mine drainages

BackgroundKlebsiella oxytoca DSM 29614 - isolated from acid mine drainages - grows anaerobically using Fe(III)-citrate as sole carbon and energy source, unlike other enterobacteria and K. oxytoca clinical isolates. The DSM 29614 strain is multi metal resistant and produces metal nanoparticles that are embedded in its very peculiar capsular exopolysaccharide. These metal nanoparticles were effective as antimicrobial and anticancer compounds, chemical catalysts and nano-fertilizers.ResultsThe DSM 29614 strain genome was sequenced and analysed by a combination of in silico procedures. Comparative genomics, performed between 85 K. oxytoca representatives and K. oxytoca DSM 29614, revealed that this bacterial group has an open pangenome, characterized by a very small core genome (1009 genes, about 2%), a high fraction of unique (43,808 genes, about 87%) and accessory genes (5559 genes, about 11%). Proteins belonging to COG categories “Carbohydrate transport and metabolism” (G), “Amino acid transport and metabolism” (E), “Coenzyme transport and metabolism” (H), “Inorganic ion transport and metabolism” (P), and “membrane biogenesis-related proteins” (M) are particularly abundant in the predicted proteome of DSM 29614 strain. The results of a protein functional enrichment analysis - based on a previous proteomic analysis – revealed metabolic optimization during Fe(III)-citrate anaerobic utilization. In this growth condition, the observed high levels of Fe(II) may be due to different flavin metal reductases and siderophores as inferred form genome analysis. The presence of genes responsible for the synthesis of exopolysaccharide and for the tolerance to heavy metals was highlighted too. The inferred genomic insights were confirmed by a set of phenotypic tests showing specific metabolic capability in terms of i) Fe2+ and exopolysaccharide production and ii) phosphatase activity involved in precipitation of metal ion-phosphate salts.ConclusionThe K. oxytoca DSM 29614 unique capabilities of using Fe(III)-citrate as sole carbon and energy source in anaerobiosis and tolerating diverse metals coincides with the presence at the genomic level of specific genes that can support i) energy metabolism optimization, ii) cell protection by the biosynthesis of a peculiar exopolysaccharide armour entrapping metal ions and iii) general and metal-specific detoxifying activities by different proteins and metabolites.

Tail tubular protein A: a dual-function tail protein of Klebsiella pneumoniae bacteriophage KP32

Tail tubular protein A (TTPA) is a structural tail protein of Klebsiella pneumoniae bacteriophage KP32, and is responsible for adhering the bacteriophage to host cells. For the first time, we found that TTPA also exhibits lytic activity towards capsular exopolysaccharide (EPS) of the multiresistant clinical strain of Klebsiella pneumoniae, PCM2713, and thus should be regarded as a dual-function macromolecule that exhibits both structural and enzymatic actions. Here, we present our crystallographic and enzymatic studies of TTPA. TTPA was crystallized and X-ray diffraction data were collected to a resolution of 1.9 Å. In the crystal, TTPA molecules were found to adopt a tetrameric structure with α-helical domains on one side and β-strands and loops on the other. The novel crystal structure of TTPA resembles those of the bacteriophage T7 tail protein gp11 and gp4 of bacteriophage P22, but TTPA contains an additional antiparallel β-sheet carrying a lectin-like domain that could be responsible for EPS binding. The enzymatic activity of TTPA may reflect the presence of a peptidoglycan hydrolase domain in the α-helical region (amino acid residues 126 to 173). These novel results provide new insights into the enzymatic mechanism through which TTPA acts on polysaccharides.

The influence of rifampicin resistant mutations on the biosynthesis of exopolysaccharides by strain Escherichia coli K-12 lon

The influence of RNA polymerase (rif) mutations on the yield of capsular exopolysaccharide--colanic acid (CA) of Escherichia coli K-12 lon strain was studied. Five colanic acid isogenic producing strains were created by transduction transfer of rif alleles possessing pleiotropic effects. The obtained isogenic strains differed by specific growth rate, size and mucoidness of colonies, the dependence of growth on the medium composition and cultivation temperature, as well as by the adsorption rate of virulent bacteriophage M59, specifically lysing E. coli cells producing CA. Direct correlation between the yield of exopolysaccharides, growth rate and adsorption of bacteriophage M59 was revealed. Among rif recombinants strain AH203, which synthesized twice as much CA compared with the parental strain in submerged cultivation was selected.

Antibiotic modulation of capsular exopolysaccharide and virulence in Acinetobacter baumannii.

Acinetobacter baumannii is an opportunistic pathogen of increasing importance due to its propensity for intractable multidrug-resistant infections in hospitals. All clinical isolates examined contain a conserved gene cluster, the K locus, which determines the production of complex polysaccharides, including an exopolysaccharide capsule known to protect against killing by host serum and to increase virulence in animal models of infection. Whether the polysaccharides determined by the K locus contribute to intrinsic defenses against antibiotics is unknown. We demonstrate here that mutants deficient in the exopolysaccharide capsule have lowered intrinsic resistance to peptide antibiotics, while a mutation affecting sugar precursors involved in both capsule and lipopolysaccharide synthesis sensitizes the bacterium to multiple antibiotic classes. We observed that, when grown in the presence of certain antibiotics below their MIC, including the translation inhibitors chloramphenicol and erythromycin, A. baumannii increases production of the K locus exopolysaccharide. Hyperproduction of capsular exopolysaccharide is reversible and non-mutational, and occurs concomitantly with increased resistance to the inducing antibiotic that is independent of the presence of the K locus. Strikingly, antibiotic-enhanced capsular exopolysaccharide production confers increased resistance to killing by host complement and increases virulence in a mouse model of systemic infection. Finally, we show that augmented capsule production upon antibiotic exposure is facilitated by transcriptional increases in K locus gene expression that are dependent on a two-component regulatory system, bfmRS. These studies reveal that the synthesis of capsule, a major pathogenicity determinant, is regulated in response to antibiotic stress. Our data are consistent with a model in which gene expression changes triggered by ineffectual antibiotic treatment cause A. baumannii to transition between states of low and high virulence potential, which may contribute to the opportunistic nature of the pathogen.

Highly dynamic genomic loci drive the synthesis of two types of capsular or secreted polysaccharides within the Mycoplasma mycoides cluster.

Mycoplasmas of the Mycoplasma mycoides cluster are all ruminant pathogens. Mycoplasma mycoides subsp. mycoides is responsible for contagious bovine pleuropneumonia and is known to produce capsular polysaccharide (CPS) and exopolysaccharide (EPS). Previous studies have strongly suggested a role for Mycoplasma mycoides subsp. mycoides polysaccharides in pathogenicity. Mycoplasma mycoides subsp. mycoides-secreted EPS was recently characterized as a β(1→6)-galactofuranose homopolymer (galactan) identical to the capsular product. Here, we extended the characterization of secreted polysaccharides to all other members of the M. mycoides cluster: M. capricolum subsp. capripneumoniae, M. capricolum subsp. capricolum, M. leachii, and M. mycoides subsp. capri (including the LC and Capri serovars). Extracted EPS was characterized by nuclear magnetic resonance, resulting in the identification of a homopolymer of β(1→2)-glucopyranose (glucan) in M. capricolum subsp. capripneumoniae and M. leachii. Monoclonal antibodies specific for this glucan and for the Mycoplasma mycoides subsp. mycoides-secreted galactan were used to detect the two polysaccharides. While M. mycoides subsp. capri strains of serovar LC produced only capsular galactan, no polysaccharide could be detected in strains of serovar Capri. All strains of M. capricolum subsp. capripneumoniae and M. leachii produced glucan CPS and EPS, whereas glucan production and localization varied among M. capricolum subsp. capricolum strains. Genes associated with polysaccharide synthesis and forming a biosynthetic pathway were predicted in all cluster members. These genes were organized in clusters within two loci representing genetic variability hot spots. Phylogenetic analysis showed that some of these genes, notably galE and glf, were acquired via horizontal gene transfer. These findings call for a reassessment of the specificity of the serological tests based on mycoplasma polysaccharides.

Identification of capsule, biofilm, lateral flagellum, and type IV pili in Vibrio mimicus strains

Vibrio mimicus is a bacterium that causes gastroenteritis; it is closely related to Vibrio cholerae, and can cause acute diarrhea like cholera- or dysentery-type diarrhea. It is distributed worldwide. Factors associated with virulence (such as hemolysins, enterotoxins, proteases, phospholipases, aerobactin, and hemagglutinin) have been identified; however, its pathogenicity mechanism is still unknown. In pathogenic Vibrio species such as V. cholerae, Vibrio. parahaemolyticus and Vibrio vulnificus, capsule, biofilms, lateral flagellum, and type IV pili are structures described as essential for pathogenicity. These structures had not been described in V. mimicus until this work. We used 20 V. mimicus strains isolated from water (6), oyster (9), and fish (5) samples and we were able to identify the capsule, biofilm, lateral flagellum, and type IV pili through phenotypic tests, electron microscopy, PCR, and sequencing. In all tested strains, we observed and identified the presence of capsular exopolysaccharide, biofilm formation in an in vitro model, as well as swarming, multiple flagellation, and pili. In addition, we identified homologous genes to those described in other bacteria of the genus in which these structures have been found. Identification of these structures in V. mimicus is a contribution to the biology of this organism and can help to reveal its pathogenic behavior.

Streptococcus thermophilus strains of plant origin as dairy starters: Isolation and characterisation

Streptococcus thermophilus strains have been isolated mainly from dairy environments. To prospect for new strains of S. thermophilus, isolation was made from different plant sources. In this study, 74 plant isolates were characterised as S. thermophilus by a polyphasic approach and by 16S rRNA sequencing. The isolates were further evaluated for their physiological and biochemical properties. Plant isolates exhibited good acid production and varying proteolytic activity. All the isolates showed acetaldehyde and capsular exopolysaccharide (EPS) production and few revealed diacetyl production. In contrast to industrial strains, six isolates were able to ferment galactose and 25 were found to have no urease activity. Both the plant isolates and reference dairy cultures were found to possess similar physiological and biochemical properties and can be considered for developing new starters.